DESCRIPTION: The use of xenogenic, costly, and complicated culture media and substrates during stem cell expansion and differentiation procedures has greatly limited their widespread use and translational relevance. To address these barriers, we have developed a synthetic nanofiber substrate that induces differentiation of mouse embryonic stem cells to neural progenitors within 1 day. Our proposed nanofibers use combinations of topography, alignment, and the tethering of up to three individually bioactive peptides where the presentation and concentration can be controlled precisely to influence cell membrane signaling interactions. Therefore, this proposal seeks to develop this multi-peptide tethered culture substrate for the directed differentiation and maturation of neurons and glia. We are uniquely positioned to develop a synthetic culture substrate capable of providing multiple functionalization sites for directed mouse embryonic stem cell differentiation; no other group to date offers more than 1 tethered peptide functionalization site. In this R15 proposal, we will direct stem cell differentiation by (i) characterizing the impact of surface-tethered bioactive molecules on neural lineage commitment (glial or neuronal) differentiation and maturity, and (ii) directing differentiation of mESCs into neural lineages via controlled surface interaction rather than soluble factors. The outcomes of these two aims will provide a novel, synthetic culture substrate for differentiation and long-term culture of mature neural cells. These outcomes have broad implications, as these culture substrates could be further modified to control the differentiation and maturity of other lineages. The PIs have excellent track records with research training, with the majority of the undergraduate students trained in their laboratories advancing into research-based, health-related graduate programs. If funded, the proposed project would significantly enhance the research environment in two colleges at The University of Akron. Both PIs have actively integrated undergraduates into their research laboratories, resulting in 18 co-authored papers (including 5 first-authored papers) over the past 5 years.